From rice bag to table: Fate of phenolic chemical compositions and antioxidant activities in waxy and non-waxy black rice during home cooking

Food Chemistry 191 (2016) 81–90

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Food Chemistry

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From rice bag to table: Fate of phenolic chemical compositions andantioxidant activities in waxy and non-waxy black rice during homecooking

http://dx.doi.org/10.1016/j.foodchem.2015.02.0010308-8146/� 2015 Elsevier Ltd. All rights reserved.

⇑ Corresponding author at: 28, Jinfeng Road, Tangjiawan, Zhuhai, Guangdong519085, China. Tel.: +86 756 3620636; fax: +86 756 3620882.

E-mail address: [email protected] (B. Xu).

Yayuan Tang, Weixi Cai, Baojun Xu ⇑Food Science and Technology Program, Beijing Normal University-Hong Kong Baptist University United International College, Zhuhai, Guangdong 519085, China

a r t i c l e i n f o

Article history:Received 13 September 2014Received in revised form 23 January 2015Accepted 1 February 2015Available online 7 February 2015

Chemical compounds studied in this article:Condensed tannin/proanthocyanidin(PubChem CID: 108065)Anthocyanin (PubChem CID: 145858)Anthocyanin 30-O-beta-D-glucoside(PubChem CID: 56928084)Cyanidin-3-glucoside (PubChem CID:197081)Peonidin-3-glucoside (PubChem CID:443654)DPPH (PubChem CID: 2735032)(+)-Catechin (PubChem CID: 9064)Gallic acid (PubChem CID: 370)Trolox (PubChem CID: 40634)

Keywords:Black riceThermal processingPhenolic compoundsAnthocyaninCyanidin-3-glucosideAntioxidants

a b s t r a c t

The objectives of this study were to systematically analyze degradation rate of functional substances,such as total phenolic content (TPC), total flavonoid content (TFC), condensed tannin content (CTC),monomeric anthocyanin content (MAC), cyanidin-3-glucoside (Cy3glc), and peonidin-3-glucoside(Pn3glc), as well as antioxidant activities in cooked waxy and non-waxy black rice through differenthome cooking manners. Results showed that greater phenolics and antioxidant capacities were detectedin non-waxy rice rather than waxy one. All processed black rice exhibited significantly (p < 0.05) lowerTPC, TFC, CTC, MAC, Cy3glc, Pn3glc, and antioxidants as compared to the raw rice. Different processingmethods significantly degraded the content and activities of antioxidants of both waxy and non-waxyblack rice. Under the same cooking time, black rice porridge retained more active substances than thatof cooked rice by rice cooker. Therefore, to maintain bioavailability of active components, black riceporridge may gain more health promoting effects.

� 2015 Elsevier Ltd. All rights reserved.

1. Introduction

Nowadays, colored rice (Oryza stiva L.) consumption is increas-ing rapidly. Both waxy and non-waxy black rice are particularlyimportantly colored rice species, and derive their names from theirnaturally purple or black pigments that have been confirmed asanthocyanins (Kong & Lee, 2010). Rice is cultivated primarily inAsian countries, such as China, Japan and Korea, and is generallyconsumed as an ingredient in refreshments by people who are

living in those countries (Tananuwong & Tewaruth, 2010). It hasbeen shown that black rice has a beneficial contribution tonutritional and therapeutic values in comparison to white rice,and these extra values make the pigmented rice important sourcesof amino acids (especially essential amino acids), vitamins andsome trace minerals (such as Fe, Zn and Cu), as well as rich naturalcolorants (Jiang, Liu, Long, & Sheng, 1999).

It has been reported that black rice might provide health benefitsto reducing risk of chronic diseases, such as cardiovascular problems,cancers (Xia et al., 2006), diabetes and its complications (Walter &Marchesan, 2011), as well as iron-deficiency anemia (Wang & Guo,2007), because of the existences of phytochemicals in black rice,such as phenolic compounds (Shen, Jin, Xiao, Lu, & Bao, 2009).

82 Y. Tang et al. / Food Chemistry 191 (2016) 81–90

Phenolics, defined as molecules with at least one aromatic ringbearing one or more hydroxyl groups, are one of the major bioac-tive substances in black rice (Shen et al., 2009). Among phenolicsubstances, flavonoids are a large subclass, containing two or morearomatic rings and each ring bearing at least one aromatic hydro-xyl and connected with a carbon bridge. Anthocyanins (one groupof water-soluble flavonoids) are the predominant pigments andfunctional phenolics in black rice. They have been proved to beessentially substantial foundations for anti-oxidation effect,because of their notably strong free-radical scavenging effects inblack rice (Chiang et al., 2006; Shen et al., 2009; Zhang, Guoet al., 2006; Zhang, Zhang et al., 2006). Five anthocyanins havebeen separated and identified. These anthocyanins are malvidin,pelargonidin-3,5-diglucoside, cyanidin-3-glucoside, cyanidin-3,5-diglucoside and peonidin-3-glucoside (Hou, Qin, Zhang, Cui, &Ren, 2013; Zhang, Guo et al., 2006; Zhang, Zhang et al., 2006).

Rice is usually cooked before its consumption, and there are twoordinarily home-cooked products in Asian countries: rice porridgeand cooked rice. A negative correlation between thermal processingand the concentration of some functional ingredients, such as phe-nolic compounds, has been proved (Walter et al., 2013). Hiemori,Koh, and Mitchell (2009) reported that all cooking processes couldgenerate a dramatic decrease in anthocyanin content of black rice.

Although a plenty of literature support the potential beneficialeffects of black rice on health, little is known about the influenceof hydrothermal cooking approaches on functional componentsand antioxidant activities in cooked black rice. The present studywas intent to evaluate functional compositions and antioxidantactivities in waxy and non-waxy black rice as affected by homecooking. The changes in total phenolic content (TPC), total flavo-noid content (TFC), condensed tannin content (CTC), monomericanthocyanin content (MAC), cyanidin-3-glucoside and peonidin-3-glucoside, as well as antioxidant capacities in waxy and non-waxy black rice with two ordinarily home-cooking methods wereinvestigated. The changes in these functional substances under dif-ferent cooking time (20, 25, 30, and 35 min) were also investigated.The consequences of this research can assist the public to under-stand in details the loss of these bioactive substances in black riceafter home cooking.

2. Materials and methods

2.1. Chemicals and reagents

Folin–Ciocalteu reagent, 2-diohenyl-1-picryhydrazyl (DPPH),(+)-catechin, and 2,4,6-tri (2-pyridyl)-s-triazine (TPTZ) were pur-chased from Shanghai Yuanye Biological Technology Co., Ltd(Shanghai, China). Anthocyanin standards of cyanidin-3-glucosideand peonidin-3-glucoside were supplied by ChemFaces Biochemi-cal Co., Ltd. (Wuhan, China). 2,6-Di-tert-butyl-p-cresol was pur-chased from Lingfeng Chemical Reagent Co., Ltd. (Shanghai,China). 6-Hydroxy-2,5,7,8-tetramethlchroman-2-carboxylic acid(Trolox) and high-performance liquid chromatography (HPLC)-grade acetonitrile was obtained from Sigma–Aldrich Co. (Shanghai,China). Vanillin was offered by Wenzhou Dongsheng ChemicalReagent Factory (Zhejiang, China). Absolute ethanol was from Tian-jin Fuyu Fine Chemical Co., Ltd. (Tianjin, China). Other chemicalreagents were supplied by Tianjin Damao Chemical Reagent Co.,Ltd (Tianjin, China). All chemicals were of analytic grade unlessspecially mentioned.

2.2. Black rice samples

The rice species used in this research was Oryza stiva L. indicaand included its waxy and non-waxy variants. Both of them werecultivated in Guilin, Guangxi Province, China.

2.3. Soaking time and determination of hydration rate

According to the method of Xu and Chang (2008a, 2008b) withslight modifications, black rice (20 g) were rinsed under runningtap water, and soaked in 100 mL of tap water at room temperaturefor 24 h. Within the initial 6 h, the weight gain of sample was mea-sured hourly as water absorption (moisture content), followed byeach 2 h for determination over the next 10 h. The last measure-ment was done at 24 h. After each weighing, samples were placedback into the soaking water. The curve about water absorption wascreated by plotting the kinetic moisture content (%) with soakingtime.

2.4. Cooking approaches and cooking time

Two thermal processes were performed for black rice. Process-ing method 1 (black rice porridge making): about 40 g of waxy andnon-waxy black rice soaked for 2.5 h was separately added with240 mL water (containing soaking water), and cooked on an elec-tric hot plate cooker. After boiling, black rice was cooked for20 min, 25 min, 30 min, and 35 min in each different sample. Pro-cessing methods 2 (cooked black rice making): presoaked (2.5 h)waxy and non-waxy (approximately 40 g) was respectively placedinto 120 mL water (containing soaking water), and cooked by anautomatic house-hold rice cooker (Luby Electronic Co., Ltd, Guang-dong, China) for 25 min. All these cooked samples were freeze-dried by freeze-drier (Labconco Corporation, Kansas City, MO,U.S.A.), and then stored in 4 �C refrigerator for further studies.

2.5. Color measurement

Color attributes of black rice samples were measured by Color-imeter (CR-410, Konica Minolta, Japan). The color was expressed inL⁄a⁄b⁄, where the L⁄ represents lightness (L⁄ = 0 yields black andL⁄ = 100 denotes white), the a⁄ expresses red (+) or green (�),and the b⁄ indicates yellow (+) or blue (�). L⁄, a⁄ and b⁄ parameterswere measured against a white background plate which weredirectly obtained from the apparatus.

2.6. Extraction of phenolics from rice samples

Accurately, 0.5 g of each ground dry sample was weighed, andextracted with 5 mL of acidic 70% acetone (acetone/water/aceticacid = 70:29.5:0.5, v/v/v) in a set of capped centrifuge tubes byshaking on an orbital shaker at ambient temperature for 3 h andsetting in the dark for 12 h. Then the extracts were centrifuged at4000 rpm for 10 min, and the supernatants were removed. Resi-dues were mixed with 5 mL of extracting solvent for the secondtime and the third time. All extracts were combined and storedat 4 �C in the dark. The extraction of every sample was conductedin triplicate.

2.7. Determination of total phenolic content (TPC)

Total phenolic content (TPC) was evaluated according to themethod of Xu and Chang (2007) with slight modifications. TPCwas demonstrated as milligrams of gallic acid equivalents (mgGAE/g black rice).

2.8. Determination of total flavonoid content (TFC)

The total flavonoid content (TFC) was determined by a colori-metric method using (+)-catechin as the standard (Xu & Chang,2007). TFC was expressed as (+)-catechin equivalents (mg CAE/gblack rice).

Fig. 1. Water absorption curves of waxy and non-waxy black rice.

Y. Tang et al. / Food Chemistry 191 (2016) 81–90 83

2.9. Determination of condensed tannin content (CTC)

The assay of condensed tannin content (CTC) was provided byXu and Chang (2007) with slight modifications. The quantity ofcondensed tannin content (CTC) was calculated as (+)-catechinequivalents (mg CAE/g black rice).

2.10. Determination of monomeric anthocyanin content (MAC)

MAC was measured based on a pH differential method describedpreviously by Lee, Durst, and Wrolstad (2005) with slight modifica-tions. The MAC was expressed as cyanidin-3 -glucoside equivalentsbecause of its historical usage for similar assays (Lee et al., 2005), aswell as occupying the predominant position in the anthocyanin ofblack rice (Zhang, Guo et al., 2006; Zhang, Zhang et al., 2006).

2.11. Determination of DPPH free radical scavenging activity

Analysis of DPPH free radical scavenging capacity was per-formed by a colorimetric method Trolox as the external standard(Xu & Chang, 2007). The results were expressed as the equivalentcontent of Trolox (lmole TE/g black rice).

2.12. Ferric reducing antioxidant power assay (FRAP)

According to the method of Xu and Chang (2007) with slightchanges, the FRAP assay was applied to determine anti-oxidativecompetence of black rice. A calibration standard curve ofFeSO4�7H2O solution was used to calculate the FRAP value withmillimoles of Fe2+ equivalents (FE) per 100 g of black rice (i.e.mmole FE/100 g black rice).

2.13. HPLC analysis of anthocyanin content

The extraction of anthocyanin was performed by modifying themethod of Xu and Chang (2008a, 2008b). The HPLC analysis wasperformed with Waters (e2695 Separations Modulek, Milford,Massachusetts, U.S.A.) equipped with Waters 2998 PhotodiodeArray Detector, using Phenomenex C18 column (250 � 4.6 mm).HPLC conditions were as follows: Solvent A, 0.1% TFA/H2O; solventB, CH3CN/H2O/TFA (50:50:0.1, v/v/v); linear gradient, initial per-centage of B (15%) to 60 min (40%); column temperature, 40 �C;flow rate, 1.0 mL/min; and injection volume, 20 lL. The detectorwas set at 520 nm.

The stock solutions of cyanidin-3-glucoside and peonidin-3-glucoside were prepared by dissolving in methanol to give a con-centration of 1.0 mg/mL. A portion of dilutions as working solutionswas 1000 lg/mL, 500 lg/mL, 200 lg/mL, 100 lg/mL, 50 lg/mL, and25 lg/mL. Calibration curves of cyanidin-3-glucoside and peonidin-3-glucoside were plotted peak areas against concentrations byduplicate injections of the six series diluted working solutions.Anthocyanin contents were expressed as milligrams of Cy3glc orPn3glc per gram of black rice (mg/g) on a dry weight basis.

2.14. Statistical analysis

All samples from different processes were extracted and testedin triplicate. The data were evaluated as mean ± standard devia-tion. The statistically significant (p < 0.05) differences were con-ducted by analysis of variance (ANOVA) applying SPSS package.

3. Results

3.1. Soaking and determination of hydration rate

In this experiment, waxy and non-waxy black rice, in triplicate,were experimented on for 24 h to measure their hydration rate, as

indicated in Fig. 1. For non-waxy black rice, its saturation point wasachieved earlier than that of waxy black rice. The non-waxy sam-ples reached a plateau after soaking for about 5 h; however, thewaxy samples spent around 7 h to enter the equilibrium phase.At saturation point, moisture content of the waxy samples(approximately 28%) was higher than that of the non-waxy sam-ples (around 24%).

3.2. Color values of raw and cooked black rice

The color values of non-processed and processed black ricewere presented in Table 1. Raw black rice samples (waxy andnon-waxy) differed in their color values. In addition, cooked riceproducts (porridge and cooked rice) from the two species pre-formed distinctly (p < 0.05) with diverse color values.

3.3. Total phenolic content (TPC) in cooked black rice

Total phenolic contents (TPC) of black rice before and afterhome-cooking processing were exhibited in Table 2. The extractsfrom different types of black rice (waxy or non-waxy) with differentcooking methods (porridge or cooked rice) performed significantdifference (p < 0.05) in their TPC values. The TPC values of waxyand non-waxy black rice before processing were 4.92 mg GAE/gand 6.97 mg GAE/g, respectively. With respect to waxy and non-waxy rice porridge, the TPC values seem to have a decreasing trendalong with the extension of cooking time. For the waxy samples, theTPC values were 4.27 mg GAE/g, 4.06 mg GAE/g, 3.53 mg GAE/g and3.37 mg GAE/g which represent boiling for 20 min, 25 min, 30 minand 35 min, respectively. Concerning the non-waxy samples, theTPC values were 6.43 mg GAE/g for 20 min, 6.10 mg GAE/g for25 min, 5.70 mg GAE/g for 30 min, and 5.30 mg GAE/g for 35 min.The TPC values of waxy and non-waxy black cooked rice by rice coo-ker were 3.79 mg GAE/g and 5.11 mg GAE/g, respectively.

3.4. Total flavonoid content (TFC) in cooked black rice

Total flavonoid contents (TFC) of the extracts from raw blackrice and processed black rice were presented in Table 2. Theextracts from two varieties differed significantly (p < 0.05) in theirTFC values. The TFC value of non-waxy rice without thermalprocessing was 3.91 mg CAE/g. For non-waxy rice porridge, fromcooking 20 min to 35 min, the TFC values declined in the rangeof 3.74–2.87 mg CAE/g. As for the cooked rice, its TFC value was3.10 mg CAE/g. The TFC value of waxy black rice before cooking

Table 1Effects of different home cooking process on color values of black rice.

Species Sample L⁄ a⁄ b⁄ DE

Waxy Raw rice 59.2 ± 0.09 a 3.19 ± 0.01 l 1.51 ± 0.02 h 35.5 ± 0.09 lPorridge, 20 min 52.9 ± 0.22 c 8.64 ± 0.07 h 3.13 ± 0.04 d 42.5 ± 0.23 JPorridge, 25 min 47.2 ± 0.31g 10.9 ± 0.01 d 4.38 ± 0.02 a 48.7 ± 0.30 ePorridge, 30 min 47.9 ± 0.19 f 10.2 ± 0.17 f 3.70 ± 0.02 c 47.8 ± 0.21 fPorridge, 35 min 48.1 ± 0.13 f 9.38 ± 0.03 g 3.93 ± 0.03 b 47.5 ± 0.13 gCooked rice, 25 min 51.2 ± 0.24 d 5.34 ± 0.02 J 2.47 ± 0.03 e 43.7 ± 0.24 i

Non-waxy Raw rice 44.8 ± 0.26h 4.54 ± 0.05 k �2.28 ± 0.04 J 50.3 ± 0.26 dPorridge, 20 min 49.0 ± 0.04 e 10.5 ± 0.08 e 1.50 ± 0.01 h 46.9 ± 0.06 hPorridge, 25 min 41.5 ± 0.03 J 13.3 ± 0.04 a 1.76 ± 0.01 g 54.8 ± 0.04 aPorridge, 30 min 43.0 ± 0.11 i 12.5 ± 0.04 c 1.74 ± 0.01 g 53.2 ± 0.12 bPorridge, 35 min 44.7 ± 0.15 h 13.0 ± 0.11 b 2.39 ± 0.02 f 51.7 ± 0.12 cCooked rice, 25 min 54.4 ± 0.08 b 8.26 ± 0.06 i -1.66 ± 0.04 i 41.4 ± 0.07 k

Data were expressed as mean ± standard deviation on dry weight basis. Values within each type of black rice marked by the same letters within same column are notsignificantly (p < 0.05) different.

Table 2Effects of different home cooking process on phenolic profiles of black rice.

Species Sample TPC (mg GAE/g) TFC (mg CAE/g) CTC (mg CAE/g) MAC (lg CyE/g)

Waxy Raw 4.92 ± 0.03 f 2.81 ± 0.12 e 11.9 ± 0.06 c 9.32 ± 0.16 gPorridge, 20 min 4.27 ± 0.09 g 2.75 ± 0.06 e 10.3 ± 0.31 ef 8.29 ± 0.19 hPorridge, 25 min 4.06 ± 0.02 g 2.50 ± 0.11 f 9.61 ± 0.10 g 7.87 ± 0.34 hPorridge, 30 min 3.53 ± 0.29 i 2.39 ± 0.06 fg 8.91 ± 0.26 h 6.52 ± 0.29 JPorridge, 35 min 3.37 ± 0.21 i 2.33 ± 0.09 g 7.36 ± 0.14 i 5.99 ± 0.06 JCooked rice, 25 min 3.79 ± 0.06 h 2.42 ± 0.05 fg 9.33 ± 0.12 g 7.20 ± 0.11 i

Non-waxy Raw 6.97 ± 0.13 a 3.91 ± 0.11 a 13.7 ± 0.10 a 22.3 ± 0.16 aPorridge, 20 min 6.43 ± 0.21 b 3.74 ± 0.06 b 12.8 ± 0.19 b 20.1 ± 0.20 bPorridge, 25 min 6.10 ± 0.18 c 3.54 ± 0.11 c 11.5 ± 0.14 d 19.2 ± 0.22 cPorridge, 30 min 5.70 ± 0.11 d 3.23 ± 0.07 d 10.0 ± 0.23 f 17.3 ± 0.62 dPorridge, 35 min 5.30 ± 0.15 e 2.87 ± 0.06 e 8.91 ± 0.08 h 15.9 ± 0.72 eCooked rice, 25 min 5.11 ± 0.05 ef 3.10 ± 0.06 d 10.5 ± 0.51 e 13.3 ± 0.24 f

Data were expressed as mean ± standard deviation on dry weight basis. Values within each type of black rice marked by the same letters within same column are notsignificantly (p < 0.05) different.

84 Y. Tang et al. / Food Chemistry 191 (2016) 81–90

was 2.81 mg CAE/g, and the cooked waxy black rice by anautomatic rice cooker was 2.42 mg CAE/g. In regard to waxy riceporridge, the TFC values were 2.75 mg CAE/g (20 min), 2.50 CAE/g(25 min), 2.39 mg CAE/g (30 min), and 2.33 CAE/g (35 min).

3.5. Condensed tannin content (CTC) in cooked rice

Condensed tannin contents (CTC) of black rice with and withoutthermal processing were presented in Table 2. The extracts fromdifferent varieties of black rice with different cooking approachesdiffered significantly (p < 0.05) in their CTC values. The CTC valuesof raw waxy and non-waxy black rice were 11.9 and 13.7 mg CAE/g, respectively. There was a downward tendency for both twotypes of black rice after processing with home cooking. For theCTC values of waxy black rice porridge, their CTC values were10.3 mg CAE/g for 20 min, 9.61 mg CAE/g for 25 min,8.91 mg CAE/g for 30 min and 7.36 mg CAE/g for 35 min. For theCTC values of non-waxy black rice porridge, their CTC values were12.8 mg CAE/g for 20 min, 11.5 mg CAE/g for 25 min, 10.0 mg CAE/g for 30 min, and 8.91 mg CAE/g for 35 min. About cooked rice byrice cooker, the thermal processing had the impacts on bothcooked waxy black rice (9.33 mg CAE/g) and cooked non-waxyblack rice (10.5 mg CAE/g).

3.6. Monomeric anthocyanin content (MAC) in cooked rice

Monomeric anthocyanin contents (MAC) of the extracts fromnon-processed and processed black rice were presented in Table 2.The black rice extracts from different thermal processing differedsignificantly (p < 0.05) in their MAC values. Meanwhile, different

species of black rice (i.e. waxy and non-waxy) exhibited variousMAC values. The MAC of processed waxy rice declined dramaticallyfrom 8.29 lg CyE/g in porridge with 20 min to 5.99 lg CyE/g inporridge with 35 min, while the raw waxy one was 9.32 lg CyE/g, and the cooked waxy rice by rice cooker displayed7.20 lg CyE/g. However, the MAC value of raw non-waxy ricewas 22.3 lg CyE/g. Along with the extension of cooking time from20 min to 35 min for porridge, the MAC varied in the range of 20.1–15.9 lg CyE/g of samples. Cooked non-waxy rice revealed13.3 lg CyE/g.

3.7. Radical DPPH scavenging activity

DPPH free radical scavenging capacities (DPPH) of the non-pro-cessed and processed black rice were listed in Table 3. Withoutthermal processing, raw black rice samples (waxy and non-waxy)differed in their DPPH values. In addition, the different kinds ofblack rice treated with two home-cooking methods preformed dis-tinctly (p < 0.05) with diverse DPPH values. The DPPH value of rawwaxy black rice was 22.7 lmole TE/g. In cooked products (eitherporridge or cooked rice), losses of DPPH values were indicated:the DPPH values of 20 min, 25 min, 30 min, and 35 min were22.2 lmole TE/g, 22.2 lmole TE/g, 21.0 lmole TE/g and 20.6 lmo-le TE/g, respectively. Meanwhile, the DPPH value for cooked waxyblack rice was 20.7 lmole TE/g. The same phenomenon was shownin non-waxy black rice which had a relatively higher DPPH value inraw sample, with 25.0 lmole TE/g. Under thermal approaches, theDPPH values of non-waxy one ranged from 24.2 lmole TE/g in por-ridge with 20 min to 22.8 lmole TE/g in porridge with 35 min, andthe cooked rice had the DPPH value at 21.6 lmole TE/g.

Table 3Effects of different home cooking process on antioxidant activities of black rice.

Species Sample DPPH(lmole TE/g)

FRAP(mmole FE/100 g)

Waxy Raw 22.7 ± 0.27 d 1.05 ± 0.02 dPorridge, 20 min 22.2 ± 0.19 e 0.91 ± 0.01 ePorridge, 25 min 22.2 ± 0.25 e 0.87 ± 0.01 efPorridge, 30 min 21.0 ± 0.07 g 0.84 ± 0.05 fgPorridge, 35 min 20.6 ± 0.13 g 0.76 ± 0.02 hCooked rice, 25 min 20.7 ± 0.42 g 0.83 ± 0.02 g

Non-waxy Raw 25.0 ± 0.35 a 1.38 ± 0.02 aPorridge, 20 min 24.2 ± 0.24 b 1.27 ± 0.01 bPorridge, 25 min 24.0 ± 0.34 b 1.20 ± 0.01 cPorridge, 30 min 23.4 ± 0.22 c 1.06 ± 0.02 dPorridge, 35 min 22.8 ± 0.05 d 1.05 ± 0.01 dCooked rice, 25 min 21.6 ± 0.21 f 1.07 ± 0.04 d

Data were expressed as mean ± standard deviation on dry weight basis. Valueswithin each type of black rice marked by the same letters within same column arenot significantly (p < 0.05) different.

Y. Tang et al. / Food Chemistry 191 (2016) 81–90 85

3.8. Ferric reducing antioxidant power assay (FRAP)

The FRAP values of the antioxidant extracts from black ricewere demonstrated in Table 3. These extracts from waxy or non-waxy black rice with different cooking methods performed signif-icant difference (p < 0.05) in their FRAP values. The FRAP value ofraw waxy black rice was 1.05 mmole FE/100 g. For the differentperiod of cooking for porridge, the FRAP values were not influencedsimilarly. Cooking within 20 min, the FRAP value was 0.91 mmo-le FE/100 g, followed by 0.87 mmole FE/100 g (25 min), 0.84 mmo-le FE/100 g (30 min), and 0.76 mmole FE/100 g (35 min). For thecooked waxy black rice, its FRAP value was 0.83 mmole FE/100 g.The FRAP value of raw non-waxy black rice was 1.38 mmole FE/100 g. Under the same condition with different treating time, forrice porridge, the FRAP values were 1.27 mmole FE/100 g withincooking 20 min, 1.20 mmole FE/100 g within cooking 25 min,1.06 mmole FE/100 g within cooking 30 min, and 1.05 mmole FE/100 g within cooking 35 min. For cooked rice, its FRAP value was1.07 mmole FE/100 g.

3.9. Anthocyanin content in cooked rice by HPLC quantification

The retention time of standard cyanidin-3-glucoside was about20.75 min (Fig. 2A), and retention time of standard peonidin-3-glu-coside was 33.67 min (Fig. 2B). Based on the analysis of HPLC, theindividual anthocyanin contents (cyanidin-3-glucoside and peoni-din-3-glucoside) of black rice before and after home cooking wereexhibited in Table 4. The extracts from waxy and non-waxy blackrice performed significant (p < 0.05) difference in their individualanthocyanin contents. Both of two thermal processing treatmentssignificantly (p < 0.05) reduced the contents of each individualanthocyanin in waxy and non-waxy black rice. In addition, underthe same processing time (25 min), non-waxy black rice porridgeretained significantly (p < 0.05) higher the contents of cyanidin-3-glucoside (Cy3glc) and peonidin-3-glucoside (Pn3glc), whereasthere was no significant differences between porridge and cookedrice made from waxy black rice.

The contents of individual anthocyanins (Cy3glc and Pn3glc)before processing were 3.30 mg Cy3glc/g and 0.70 mg Pn3glc/g inwaxy rice, 10.85 mg Cy3glc/g and 3.62 mg Pn3glc/g in non-waxyrice, respectively. Anthocyanins in cooked rice by rice cooker were1.13 mg Cy3glc/g and 2.04 mg Cy3glc/g in waxy rice, and 0.26 mgPn3glc/g and 0.59 mg Pn3glc/g in non-waxy rice, respectively.

With respect to non-waxy rice porridge, the anthocyanin valuesseem to have the decreasing trends along with the extension ofcooking time. For non-waxy black rice, the Cy3glc values were

5.16 mg/g for 20 min, 4.88 mg/g for 25 min, 4.61 mg/g for 30 minand 3.42 mg/g for 35 min. The same tendency was shown for thecontents of Pn3glc, decreasing from 1.64 mg/g to 1.16 mg/g. How-ever, for waxy black rice, there was no significant difference amongthe results: the Cy3glc values were 1.30 mg/g, 0.98 mg/g, 0.93 mg/g and 0.67 mg/g which represent cooking within 20 min, 25 min,30 min, and 35 min. The contents of peonidin-3-glucoside were0.28 mg/g, 0.23 mg/g, 0.23 mg/g and 0.17 mg/g accompanying bythe increase of cooking time (20 min, 25 min, 30 min and 35 min).

4. Discussion

4.1. Soaking time and water absorption

For traditional home-cooking methods in China, black rice issoaked prior to cooking, in order to decrease cooking time. In thisexperiment, black waxy rice and black non-waxy rice were soakedin triplicate for 24 h to dynamically measure their hydration rate.

Generally, both samples are black rice, yet they have dissimilarwater absorption behaviors, because of the different varieties. Forwaxy black rice, its saturation point was reached later than thatof non-waxy black rice. Non-waxy rice entered the equilibriumphase after soaking for about 5 h; however, waxy rice spent around7 h to achieve saturation. In addition, after soaking, the water pre-sents the color purple, which means plenty of water-soluble sub-stances were dissolved (possibly include phenolic antioxidants). Itcan be observed that the water for soaking non-waxy rice had adeeper violet than the soaking water used for waxy rice. Therefore,based on this phenomenon, it seems that the amount of pigment,mainly anthocyanin, in non-waxy rice is much larger than that inwaxy black rice. According to the experiments of TPC, TFC andMAC in this study, it can be phenolic contents were higher in rawnon-waxy black rice (6.97 mg GAE/g for TPC, 3.91 mg CAE/g forTFC, 22.3 lg CyE/g for MAC), rather than raw waxy black rice(4.92 mg GAE/g for TPC, 2.81 mg CAE/g for TFC, 9.32 lg CyE/g forMAC). These data can be seen in Table 2.

The soaking time in this experiment was chosen 2.5 h as for bothblack non-waxy rice and black waxy rice. The reason for choosing2.5 h as soaking period was to follow the ordinary home-cookingapproaches which are mentioned by many recipes and to decreasethe potential loss of these water-soluble constituents.

4.2. Color values of cooked rice

In Table 1, the color of black rice was significantly affected bythermal processing, expressing as L*a*b*. It can be seen clearly thatunder heat treatment, the color of waxy black rice and non-waxyblack rice had been changed. However, the change did not presenta linear tendency with the increasing of cooking time. The reasonmay be the existences of several types of individual anthocyanin,such as malvidin, pelargonidin-3,5-diglucoside, cyanidin-3-gluco-side and cyanidin-3,5-diglucoside (Zhang, Guo et al., 2006;Zhang, Zhang et al., 2006). Within the different time, the degrada-tion rate of each individual anthocyanin is different, which can leadto different color values.

4.3. Effect of processing on phenolic chemical compositions in blackrice

Phenolic substances are secondary metabolites of plants, withvarious functions such as protection against pathogens and preda-tors, mechanical support, attraction of pollinating animals, andprevention of ultraviolet radiation (Walter & Marchesan, 2011).Condensed tannins, also named proanthocyanidins (PAs), consistof polymerized flavanol subunits (Schofield, Mbugua, & Pell,

Fig. 2. HPLC chromatograms of (A) cyanidin-3-glucoside standard (500 lg/mL), (B) peonidin-3-glucoside standard (500 lg/mL), (C) raw black waxy rice, and (D) raw blacknon-waxy rice at 520 nm.

86 Y. Tang et al. / Food Chemistry 191 (2016) 81–90

Table 4Effects of different home cooking process on anthocyanin contents of black rice.

Species Sample Cyanidin-3-glucoside(mg/g)

Peonidin-3-glucoside(mg/g)

Waxy Raw 3.30 ± 0.43 c 0.70 ± 0.11 dPorridge, 20 min 1.30 ± 0.16 de 0.28 ± 0.03 ePorridge, 25 min 0.98 ± 0.03 e 0.23 ± 0.01 ePorridge, 30 min 0.93 ± 0.13 e 0.23 ± 0.04 ePorridge, 35 min 0.67 ± 0.06 e 0.17 ± 0.02 eCooked rice, 25 min 1.13 ± 0.36 de 0.26 ± 0.06 e

Non-waxy Raw 10.9 ± 0.92 a 3.62 ± 0.09 aPorridge, 20 min 5.16 ± 0.18 b 1.64 ± 0.01 bPorridge, 25 min 4.88 ± 0.61 b 1.54 ± 0.09 bPorridge, 30 min 4.61 ± 0.23 b 1.52 ± 0.08 bPorridge, 35 min 3.42 ± 0.23 c 1.16 ± 0.08 cCooked rice, 25 min 2.04 ± 0.81 d 0.59 ± 0.40 d

Data were expressed as mean ± standard deviation on dry weight basis. Valueswithin each type of black rice marked by the same letter within same column arenot significantly (p < 0.05) different.

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2001). Proanthocyanidins can yield anthocyanidins, when depoly-merization reacts under oxidative conditions. It contains higherantioxidant activity in vitro, compared to monomeric phenolicsubstances (Dyekes & Rooney, 2007). Meanwhile, condensed tan-nins can have the protective and modulating properties of cancer,cardiovascular diseases, gastrointestinal diseases, and cholesterollevel (de la Iglesia, Milagro, Campión, Boqué, & Martínez, 2010).

The phenolic substances in the rice extracts are determined col-orimetrically using Folin–Ciocalteu. The values of raw black rice, yetvarious literature presented different TPC values of raw black rice.For instance, Moongngarma, Daomukdaa, and Khumpika (2012)reported that waxy black rice contained 7.14 mg GAE/g in branlayer and 6.65 mg GAE/g in rice (without bran layer); Yodmanee,Karrila, and Pakdeechanuan (2011) reported that 2.80 mg GAE/gand 3.29 mg GAE/g for two different types of black rice. However,for this research, the TPC values of waxy and non-waxy black ricewere 4.92 mg GAE/g and 6.97 mg GAE/g, respectively.

Possibly, black rice growing under different environments andregions produce different physical characteristics and chemicalcompositions. Black rice samples in the current study were fromGuilin in Guangxi Province, therefore, variations of the TPC, TFCand CTC values were found within the rice varieties. Apart from thisreason, the method of extraction plays an important role. In thisresearch, black rice was extracted twice with 5 mL of acidic 70%acetone (acetone/water/acetic acid = 70:29.5:0.5, v/v/v). However,for some previous studies, the determination of TPC by RajaSulaiman, Osman, Saari, and Abdul-Hamid (2007) used 80% metha-nol containing 1% HCl as the extracting solution; Tananuwong andTewaruth (2010) used acetone: water mixture (70:30, v/v) with dif-ferent pH values (pH 2 and 6.8), so that they obtained different TPCvalues for the black rice with the same variety. In addition, the TPC,TFC and CTC values of waxy rice was lower than that of non-waxyrice agree with the phenomenon in determination of waterhydration, in which was the darker purple in the soaking water ofnon-waxy black rice, rather than that of waxy one.

The thermal processing can lead to partial loss of phenolics inblack rice (Shen et al., 2009). This study can give support on thisviewpoint. Compared to the data of the raw rice, the TPC, TFCand CTC values of waxy and non-waxy black rice were affectedby home-cooking methods either for porridge or for cooked rice.It can be seen clearly that the TPC values were declined accompa-nying by the extension of cooking time (Table 2). For example, non-waxy black rice porridge had 6.43 mg GAE/g for 20 min, and5.30 mg GAE/g for 35 min. The similar tendency of consequenceswas also shown in the TFC and CTC values. For example, CTC valuewas 11.9 mg CAE/g in raw waxy black rice, however, after thermal

processing, it had 10.3 mg CAE/g in waxy black rice porridge withincooking 20 min, and 7.36 mg CAE/g for 35 min cooking period.There have been several relevant findings about the effect of ther-mal processing on the content of tannin, for example, Makkar andBecker (1996) reported that the recovery of tannins decreased withthe increases in storage time, pH, and temperature.

Although these two thermal processing exhibited a negativeeffect on TPC, the degree of influence was different. The cooked ricewaxy and non-waxy rice porridge differed significantly (p < 0.05)in their TPC values. Under the similar processing time, cookingblack rice by an automatic rice cooker can produce much moresevere influence on the TPC than porridge cooking by the electrichot plate cooker. In waxy black rice, within 25 min cooking time,the data from the two thermal processing methods had no signifi-cant difference in their TFC values. Meanwhile, there was also noconsiderable difference in their CTC values. However, for non-waxyone, significant (p < 0.05) differences in both TFC values and CTCvalues existed between the two cooking treatments.

4.4. Effect of processing on anthocyanin compositions in black rice

Anthocyanin as one of the main flavonoids has a responsibilityfor the attractive purple color in black rice, and it accumulatesmainly in rice bran during maturation (Mónica Giusti &Worlstad, 2001). The amount of anthocyanin presents in a materialby measuring the color change in absorbance at two different pHvalues, where the colored form exists at pH 1.0 and the colorlessform predominates at pH 4.5 (Lee et al., 2005; Sondheimer &Kertesz, 1948), because of a reversible structural transformationof anthocyanins as a function of pH (Lee et al., 2005). Results areexpressed as cyanidin-3-glucoside (Cy3glc) equivalents, which isthe most common type of anthocyanin pigments found in nature(Lee et al., 2005).

Monomeric anthocyanin content (MAC) of the extracts fromnon-processed and processed black rice is compared distinctly.The MAC of unprocessed black non-waxy rice (22.3 lg CyE/g)was much higher than that of raw waxy rice (9.32 lg CyE/g). Thisconsequence is satisfied with the phenomenon in the determina-tion of water absorption, which exhibited that the violet color oftap water for soaking waxy black rice was deeper than that fornon-waxy black rice.

After thermal processing, the MAC values of waxy and non-waxy black rice were decreased significantly, and with the exten-sion of cooking period, the loss of anthocyanin content in sampleswas increased. For example, there was a downward trend in therange of cooking period within 20 min and 35 min for waxy blackrice porridge, the change of MAC values was from 8.28 lg CyE/g to5.99 lg CyE/g. Anthocyanins are relatively unstable and oftenundergo degradative reactions under unfavorable conditions, suchas high temperature and light (Xu & Chang, 2009). Thermal pro-cessing might cause degradation of polyphenols and release boundphenolic compositions (Xu & Chang, 2009).

In addition, both waxy and non-waxy rice were seriouslyaffected by cooking in an automatic rice cooker as cooked rice.Comparing with the data of rice as porridge, cooked rice (eitherwaxy or non-waxy rice) exhibited significant (p < 0.05) decreasesin the MAC values. During the cooking period (25 min), cookedwaxy rice (7.20 lg CyE/g) and non-waxy rice (13.4 lg CyE/g)reduced more MAC values than that of rice porridge within25 min (7.87 lg CyE/g for waxy one and 19.2 lg CyE/g for non-waxy one). The temperature and pressure of the automatic ricecooker might be higher than that of the electric hot plate cooker,therefore, even though under the same period (25 min) for cook-ing, the degree of degradation on anthocyanins, which are verysensitive to temperature, was different.

88 Y. Tang et al. / Food Chemistry 191 (2016) 81–90

Anthocyanin determination based on pH differential methodwith color reaction may sometimes overestimate anthocyanincontent, because of certain non-anthocyanin substances giving apositive reaction (Xu & Chang, 2009). Hence, anthocyanin contentfor identification and quantification was further detected usingHPLC analysis. However, HPLC cannot quantify all anthocyaninsunder the specific wavelength. This is the reason why there shouldbe multiple methods to determine anthocyanin content by combi-nation of colorimetric method by UV–Vis spectroscopy and HPLCmethods in this research.

Numerous studies have suggested that cyanidin-3-glucoside isthe most dominant anthocyanin, and the minor one is peonidin-3-glucoside (Hou et al., 2013; Kim et al., 2008; Zhang, Guo et al.,2006; Zhang, Zhang et al., 2006). However, the composition andcontents of anthocyanins are different in black rice, because antho-cyanins are not stable and vulnerable to be destroyed by a numberof factors, such as pH values, light, and thermal treatments (Houet al., 2013). Oxygen also plays a pivotal role in the destructionof anthocyanins, because oxygen as the medium can make poly-phenol oxidase catalyze indirectly the color deterioration and theloss of anthocyanins (Howard, Brownmiller, & Prior, 2014). Oxygenexclusion and samples prepared under carbon dioxide or nitrogenmay improve color and anthocyanin stability (Howard et al., 2014).In this research, two anthocyanins (cyanidin-3-glucoside andpeonidin-3-glucoside) were analyzed in waxy and non-waxy blackrice with or without thermal processing.

According to Kim et al. (2010), the retention time of cyanidin-3-glucoside (Cy3glc) was shorter than that of peonidin-3-glucoside(Pn3glc), which has been reported by Jang et al. (2012) as well. Thiscan be proved in current study, where was shown about 21 min asthe retention time of Cy3glc and around 34 min for detection ofPn3glc. Non-processed waxy and non-waxy black rice differed sig-nificantly (p < 0.05) in their individual anthocyanin content (shownin Table 4). The raw black waxy rice contained cyanidin-3-gluco-side with 3.30 mg/g and peonidin-3-glucoside with 0.70 mg/g.However, the non-waxy black rice had Cy3glc 10.9 mg/g andPn3glc 3.62 mg/g, which were much higher than those in waxyone. Therefore, these results can conform to the phenomenon inthe determination of soaking time that the deeper purple colorwas observed for non-waxy black rice. It is also satisfied with theintensity of oxidation resistance, reflecting in the experiments ofantioxidant activities (DPPH and FRAP).

Commonly, there are two home-cooked products (rice porridgeand cooked rice) in China. In comparison to black rice without heatprocessing (raw samples), two thermal methods exerted the nega-tive effects on the individual anthocyanin contents (Cy3glc andPn3glc) of waxy and non-waxy black rice. For example, in waxyblack rice, the content of cyanidin-3-glucoside was 3.30 mg/g,and concerning waxy black rice porridge, the amount of Cy3glcchanged from 1.30 mg/g to 0.67 mg/g. According to Hou et al.(2013), anthocyanins are unstable to high temperature, particu-larly for high pH conditions. Therefore, it is important to controlthe temperature, since the rate of degradation of anthocyaninsincreases as temperature rises (Escribano-Bailón, Santos-Buelga,& Rivas-Gonzalo, 2004).

In details, discussing about the individual anthocyanins, exceptthe Pn3glc content in waxy black rice, the content of Cy3glc inwaxy black rice and non-waxy black rice, as well as the contentof Pn3glc in non-waxy black rice were influenced by the timechange during heat processing. In other words, during heat pro-cessing for rice porridge, the time of waxy rice sample did notaffect the Pn3glc content under this statistical analysis. The Pn3glccontent in raw waxy black rice was only 0.70 mg/g which was theminimum amount in the individual anthocyanin. After thermaldegradation of anthocyanin, the content of peonidin reduced alot. Therefore, the significant differences in the peonidin contents

under the influence of time change might become smaller, or itmight be difficult to detect the differences from the vanishinglysmall amounts under the current technical conditions.

In relation to the stability of anthocyanins, its stability increaseswith the number of methoxyls in the B ring and decreases ashydroxyls increase, therefore, the more stable substance waspeonidin (substitution pattern: 3,5,7,4-OH; 3-OMe) resistant tothermal processing, rather than cyanidin (substitution pattern:3,5,7,3,4-OH) (Escribano-Bailón et al., 2004). Peonidin-3-glucosidewas lost completely in black soybean after during thermal process-ing (Xu & Chang, 2008a; Xu & Chang, 2008b). For this study, theaverage degradation rates of cyanidins and peonidins were about73.5% and 72.5%, respectively. There was not a noticeable differ-ence between degradation rates. The results could not reflect andagree the conclusion from neither Escribano-Bailón et al. (2004)nor Xu and Chang (2008a), Xu and Chang (2008b). The degradationrates of both individual anthocyanins were nearly the same.

Both Cy3glc and Pn3glc contents in non-waxy black rice dif-fered significantly (p < 0.05) between rice porridge and cooked rice.Within the same cooking time (25 min), porridge can maintainmuch more anthocyanin content. In other words, it had strongerreducing ability. Temperature might be the influence factor. Thetemperature of the automatic rice cooker might be higher than thatof the electric hot plate cooker, therefore, even though under thesame period (25 min) for cooking, the degree of degradation onanthocyanins, which are very sensitive to temperature, was differ-ent. However, in relation to waxy rice, there were no significantdifferences between rice as porridge and cooked rice.

4.5. Effects of thermal processing on antioxidant activities of cookedblack rice

Recently, phenolic substances have been proved that couldexert their protective functions, rather than mere antioxidantproperties. Polyphenols can play an important role in treatingchronic metabolic syndrome and other some chronic diseases(e.g. cardiovascular disease and hyperlipidemia), by means ofincreasing gene transcription of Nrf2 (nuclear factor-erythroid 2-related factor 2) (Chiva-Blanch & Visioli, 2012), stimulating theexpression of PPARs (peroxisome proliferator-activated receptors),and activating the AMPk/SIRT1 (50 adenosine monophosphate-acti-vated protein kinase/sirtuin 1) signaling cascade (Giampieri,Alvarez-Suarez, & Battino, 2014). These activities can exert differ-ent biological benefits, including the regulation of antioxidantfunctions and detoxification (Chiva-Blanch & Visioli, 2012), thedecrease of risk factors for type II diabetes and metabolic disorders(e.g. fatty acid oxidation and lipid metabolism, insulin sensitivity,glucose homeostasis), and an increase in cellular metabolism(Giampieri et al., 2014). In addition, polyphenols act as modulatorsof cellular processes for cancer prevention and treatment beyondtheir antioxidant capacity (Giampieri et al., 2014). Phenols can reg-ulate cell signaling in the tumor cells for inhibiting proliferation ofthe cells, by means of demethylation of tumor suppressor genes,inducing cell cycle arrest, and suppressing tumor angiogenesis(Giampieri et al., 2014). The ultimate purpose is to make apoptosisof cancer cells. Furthermore, there is the development of the bidi-rectional benefits between polyphenols and the microbiota (e.g.Firmicutes and Bacteroidetes) in the intestine (Chiva-Blanch &Visioli, 2012).

Although some new findings have supported polyphenols’ func-tions besides antioxidation, their antioxidant capacity (scavengingROS/RNS or limiting their formation) is still the most famous andacceptable mechanism related to health. For example, antioxidantcapacity is the major mechanism in polyphenols’ anti-carcinogen-esis effectiveness and the suppression of cancer initiation(Giampieri et al., 2014). Therefore, the experiments and discussion

Y. Tang et al. / Food Chemistry 191 (2016) 81–90 89

will be concentrated on polyphenols serving as antioxidants. Forthis article, two methods for assessment were conducted: DPPHand FRAP.

The anti-oxidative efficiency of components is indicated by thedegree of discoloration of the violet solution containing DPPHwhich is a free radical and stable at room temperature (Xu &Chang, 2007). DPPH is the nitrogen-centered radical, and themethod is mainly used for lipophilic antioxidants (Niki, 2011).DPPH free radical scavenging capacities (DPPH) of the non-pro-cessed and processed black rice were mentioned in Table 3. TheDPPH value of non-processed (i.e. raw) non-waxy black rice(25.0 lmole TE/g) was greater than that of waxy black rice(22.7 lmole TE/g). Therefore, non-waxy black rice had much moreantioxidant properties. With home-cooking methods, both twovarieties (non-waxy and waxy) exhibited a decreasing tendency,along with the increasing of time. However, different thermalprocessing methods can trigger significant diversity of the DPPHvalues.

In comparison, in terms of both two types of black rice (waxyand non-waxy), the thermal process for cooked rice affected moreon DPPH radical scavenging capacity of black rice. The rice cookedby an automatic rice cooker was treated for about 25 min, whichcan compare with the porridge of 25 min. Under the same cookingperiod, the DPPH value was higher in rice porridge (22.2 lmole TE/g for waxy one and 24.0 lmole TE/g for non-waxy one), rather thancooked rice (20.7 lmole TE/g for waxy one and 21.6 lmole TE/g fornon-waxy one). Consequently, porridge within cooking 25 mincan maintain much more DPPH radical scavenging activity ofantioxidants.

The specificity and sensitivity of one method does not providethe complete findings of all phenolic components in the extracts(Xu & Chang, 2009). Hence, a combination of two tests could sup-ply a more reliable assessment of the antioxidant properties ofwaxy and non-waxy black rice. FRAP applies the electron-transfermechanism for the determination of antioxidant activity (Prior,Wu, & Schaich, 2005). The principle of FRAP as say is that with areductant (antioxidants) at low pH, ferric tripyridyltriazine(Fe(III)-TPTZ) is reduced to the ferrous tripyridyltriazine (Fe(II)-TPTZ) that has an intensive blue color and can be detected at thewavelength of 593 nm (Benzie & Straint, 1996; Xu & Chang, 2007).

The FRAP values of raw waxy and non-waxy black rice weremeasured individually with 1.05 mmole FE/100 g and 1.38 mmo-le FE/100 g in the current study. Hence, relatively stronger antiox-idant activity profiles were shown in non-waxy black rice, ratherthan waxy rice. A similar result had been also gained from thedetermination of radical DPPH scavenging activity. However, therewere a plenty of previous documents about the FRAP values ofblack rice without thermal processing. For example, Sompong,Siebenhandl-Ehn, Linsberger-Martin, & Berghofer, 2011 reported5.50 mmole FE/100 g as the FRAP value of black rice.

As with the influence factors for the DPPH values, the FRAP val-ues of black rice were changed by thermal processing. With twohome-cooking methods (one for porridge and the other for cookedrice), there was an obvious decrease in the FRAP values of pro-cessed black rice, compared to non-processed rice. Significant dif-ferences (p < 0.05) in FRAP values were found before and afterprocessing treatments for both waxy and non-waxy rice. For exam-ple, concerning waxy black rice, its FRAP values reflected 0.91, 0.87,0.84, and 0.76 mmole FE/100 g in every interval of 5 min (from20 min to 35 min). The destruction of antioxidant compounds(e.g. phenolic substances) by boiling might be the reason to explainthe reducing trend of the FRAP values (Xu & Chang, 2009).

In comparison to the heat treatment of black rice porridge,within the same cooking time, cooked rice by an automatic ricecooker retained significantly (p < 0.05) higher FRAP value. The datawere demonstrated in Table 3. For example, with respect to non-

waxy black rice, 1.20 mmole FE/100 g was the FRAP value for therice porridge cooked for 25 min, but the lower amount of FRAPvalue was detected, when non-waxy rice was cooked as cookedrice by the automatic rice cooker, its FRAP was 1.07 mmole FE/100 g. Therefore, black rice porridge preserved greater antioxidantvalues.

Some literatures have shown many different relationshipsbetween total antioxidant activities and total phenolic content(TPC). Possibly, a negative correlation between antioxidant capac-ity and total phenolic content was confirmed in strawberry species,and a positive relation was reported in peaches, as well as the cor-relation was independent in apricots (Scalzo, Politi, Pellegrini,Mezzetti, & Battino, 2005). For this study, all the above resultsfor black rice (Table 3.) reflected the same trend in total phenoliccontent (TPC), which can supply support to the assumption thatmay be the structural change of phenolic. Thermal processingexhibited a moderate negative effect on the capacity of reductionof metal ion in FRAP evaluation.

According to previous literature (Niki, 2011), black rice hasplenty of phenolic substances and other kinds of materials contrib-ute to ‘‘antioxidant capacity’’. However, the term ‘‘antioxidantcapacity’’ has many different meanings under different occasions,such as capacity for scavenging free radical, capacity for inhibitionof oxidation, or capacity for prevention of disease. Therefore, it isimportant to specify ‘‘antioxidant capacity’’ correlating with partic-ular assays. In other words, there is the lack of correlation betweendifferent methods for the same antioxidants (Niki, 2011). DPPH wasmeasured by the reaction of the sample substances with referencefree radical (DPPH solution) for the capacity of scavenging free rad-icals, whereas FRAP was assessed by stabilizing free radicals fromdonating electron for the capacity of reduction of metal ions (ferricion Fe3+) (Niki, 2011). Hence, there should be no comparisonbetween DPPH and FRAP assessment.

5. Conclusions

Thermal treatment could reduce nutritional values of black riceby chemical reactions. In current research, the negative relationshipbetween cooking period and contents of bioactive substances(including TPC, TFC, CTC, anthocyanin content and antioxidants),were investigated. The results indicated that through heat treat-ment, it can decrease these bioactive compounds significantly(p < 0.05). In addition, two thermal processing triggered differentvalues of those functional compounds. In details, black rice porridgecan retain much more antioxidants (such as phenolics). Therefore,to maintain bioavailability of active components, rice porridge asthe form of cooking black rice may gain more health promotingeffects. Black rice is always needed to cook before consumption.In other words, we cannot obtain all bioactive compounds in rawblack rice, because of thermal degradation of phenolic substances.

Acknowledgements

This research was supported by two research grants (UICRG201329, UICRG 201402) from Beijing Normal University-HongKong Baptist University United International College, China. Theauthors would like to thank Miss Chelsea Grace O’ Hara for improv-ing English language.

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